Novel penicillins and their preparation

ABSTRACT

Novel penicillins of the formula: ##STR1## wherein the ring A is a benzene ring or a 5 or 6-membered heteroaromatic ring containing one or two nitrogen atoms as the hetero atom, on which one or more of lower alkyl, lower alkoxy, lower alkylthio, lower haloalkyl, lower alkylenedioxy halogen, hydroxyl, nitro, free or protected amino, lower alkylamino, di(lower)alkylamino and lower alkanoylamino may be present, Z is a nitrogen atom or a methylene group, X is an oxygen atom or a sulfur atom, Y is a hydrogen atom, a lower alkoxycarbonyl group or a lower alkanoyl group, R 1  is a free or protected hydroxyl group and R 2  and R 3  are each a hydrogen atom or a halogen atom, and their non-toxic pharmaceutically acceptable salts, and their preparation.

This application is a division of copending application Ser. No.424,271, filed on Dec. 13, 1973 now U.S. Pat. No. 3,954,753.

The present invention relates to novel penicillins and theirpreparation. More particularly, it relates to novel penicillins andtheir non-toxic, pharmaceutically acceptable salts, which are useful asantimicrobial agents having a broad antimicrobial spectrum includingPseudomonas, and to their preparation.

It is well known that 6-(α-aminoacylamido)penicillanic acid derivativessuch as 6-(α-amino-phenylacetamido)penicillanic acid (Ampicillin),6-(α-amino-p-hydroxyphenylacetamido)penicillanic acid (Amoxycillin),6-(α-amino-thienylacetamido)penicillanic acid,6-(1-amino-cyclohexane)carboxamidopenicillanic acid (Cyclacillin), 6-(α-amino-isothiazolylacetamido)penicillanic acid and6-[α-amino-2-(1,4-hexadienylacetamido)]penicillanic acid (Epicillin)inhibit the growth of various gram-positive and gram-negative bacteria.Particularly, ampicillin is one of the excellent chemotherapeutics.These compounds, however, do not exert any appreciable antimicrobialactivity against Pseudomonas. In U.S. Pat. No. 3,433,784, there aredescribed some N-acyl derivatives of ampicillin as showing a minimalinhibitory concentration of 125 to 250 μg/ml against Pseudomonaspyocinea A or R 59, when determined by the standard test method. Theanit-Pseudomonas activity of the compounds as described in the workingexamples is, however, not so high and the antimicrobial activity againstother gram-positive and gram-negative bacteria is considerably low.Thus, it may be said that the N-acyl derivatives of ampicillin are lessvaluable than ampicillin itself from the practical viewpoint.

As the result of the study seeking novel penicillins which have a broadantimicrobial spectrum and are highly active against gram-positive andgram-negative bacteria including Pseudomonas, it has been found that,among various compounds, the penicillins of the following formulacharacteristically exhibit a noticeable antimicrobial activity againstPseudomonas and a broad antimicrobial spectrum: ##STR2## wherein thering A is a benzene ring or a 5 or 6-membered heteroaromatic ringcontaining one or two nitrogen atoms as the hetero atom, on which one ormore of lower alkyl, lower alkoxy, lower alkylthio, lower haloalkyl,lower alkylenedioxy, halogen, hydroxyl, nitro, free or protected amino,lower alkylamino, di(lower)alkylamino and lower alkanoylamino may bepresent, Z is a nitrogen atom or a methylene group, X is an oxygen atomor a sulfur atom, Y is a hydrogen atom, a lower alkoxycarbonyl group ora lower alkanoyl group, R₁ is a free or protected hydroxyl group and R₂and R₃ are each a hydrogen atom or a halogen atom.

Accordingly, a main object of the present invention is to provide novelpenicillins (I) and their non-toxic salts, which are useful asantimicrobial agents. Another object of this invention is to provide aprocess for preparing the penicillins (I) and their non-toxic salts. Afurther object of the invention is to provide a use of the penicillins(I) and their non-toxic salts as antimicrobial agents. These and otherobjects of the invention will be apparent to those conversant with theart from the foregoing and subsequent descriptions.

As to the significances of the symbols in the said formula (I) and inany other formula as hereinafter shown, the term "lower alkyl" isintended to mean generally both straight and branched chain aliphatichydrocarbon groups having not more than eight carbon atoms (preferablynot more than five carbon atoms) such as methyl, ethyl, propyl,isopropyl, butyl, isobutyl, amyl and isoamyl. Similarly where the term"lower" is used as a part of the description of any other group (e.g.lower alkoxy, lower alkylthio, lower haloalkyl, lower alkylamino,di(lower)alkylamino), it refers to the alkyl portion of such group.Thus, the terms "lower alkanoyl" and "lower alkoxycarbonyl" meansrespectively alkanoyl and alkoxycarbonyl having not more than ninecarbon atoms (preferably not more than six carbon atoms). Exceptionally,however, the term "lower alkylenedioxy" indicates alkylenedioxy havingnot more than three carbon atoms. The halogen atom includes the chlorineatom, bromine atom, iodine atom and fluorine atom.

The protected amino group and the protected hydroxyl group indicaterespectively an amino group and a hydroxyl group which are protected byany protective group conventionally employed for the protection of aminoor hydroxyl. Examples of the protected amino group include loweralkanoylamino and lower alkoxycarbonylamino. Examples of the protectedhydroxyl group include lower alkanoyloxy and lower alkoxycarbonyloxy.

The 5 or 6-membered heteroaromatic ring containing one or two nitrogenatoms may be, for example, a pyrazole ring, a thiazole ring, animidazole ring, a pyridine ring, a pyrazine ring, a pyridazine ring, apyrimidine ring, etc. Among them, preferred are a pyrazole ring, athiazole ring, a pyrazine ring and a pyrimidine ring, particularly apyridine ring.

The non-toxic, pharmaceutically acceptable salts of the penicillins (I)are, for instance, the alkali metal salts (e.g. sodium, potassiumsalts), the alkaline earth metal salts (e.g. calcium, magnesium salts),the arginine salt, the substituted and unsubstituted ammonium salts,etc. Examples of the substituted ammonium salts include the salts oftriethylamine, procaine, dibenzylamine, N-benzyl-β-phenethylamine,1-ephenamine, N,N'-dibenzylethylenediamine, dehydroabiethylamine,N,N'-bis-dehydroabiethylethylenediamine, etc.

One of the structural characteristics of the penicillins (I) of theinvention is that the residue ##STR3## bears thereon the substituent--X--Y linked to a carbon atom adjacent to the carbon atom to which a6-(α-aminoacylamido)penicillanic acid moiety is linked. The compoundswherein the residue ##STR4## bears no such substituent areantimicrobially much less active than those bearing the substituent andexhibit only the same low antimicrobial activity as those disclosed inU.S. Pat. No. 3,433,784 against Pseudomonas as well as othergram-positive and gram-negative bacteria.

Another structural characteristic of the penicillins (I) is the presenceof the substituent R₁. The serum and urinary concentrations in mice andrats of the compounds wherein the phenyl group bears the substituent R₁are higher than those of the compounds not having such substituent.

According to the present invention, the penicillin (I) can be producedby reacting a carboxylic acid of the formula: ##STR5## wherein A, Z, X,Y, R₁, R₂ and R₃ are each as defined above and n is 0 or 1, or itsreactive derivative with an amine of the formula: ##STR6## wherein R₁,R₂, R₃ and n are each as defined above, or its derivative, if necessary,followed by hydrolysis, reduction or acylation of the resulting productand/or elimination of any protective group.

The reaction can be carried out in a conventional coupling method and/orby the use of a conventional coupling reagent in the related art field,i.e. in the synthesis of peptides, penicillins, cephalosphorins and thelike.

The compound (II) may be used as such, i.e. in a free or salt form, oras the reactive derivative. Examples of salts of the compound (II) arethe salts of alkali metals, alkaline earth metals, ammonia and organicbases (e.g. trimethylamine, triethylamine, dicyclohexylamine).

The reactive derivatives of the compound (II) on the carboxyl groupinclude, e.g. acid halides, acid anhydrides, active amides, acid azidesand active esters. Among the acid halides, the use of an acid chlorideis the most favorable. Examples of the acid anhydrides are mixed acidanhydrides and symmetric acid anhydrides prepared by the use of acidssuch as toluenesulfonic acid, an alkylcarbonic acid and an aliphaticcarboxylic acid (e.g. pivalic acid). Examples of the active amides arethose obtained by using imidazole, dimethylpyrazole, triazole, tetrazoleor the like. Examples of the active esters are those prepared by usingp-nitrophenol, pentachlorophenol, p-nitrothiophenol,N,N'-dimethylhydroxylamine, 1-hydroxy-2(1H)-pyridone,N-hydroxysuccinimide or N-hydroxyphthalimide.

When the compound (II) wherein Y is hydrogen or its reactive derivativeis used, the hydroxyl group may be protected with any protective groupas is conventionally employed in the related art field.

Illustrating some of the reactive derivatives of the compound (II)wherein n is 0 in detail, the mixed acid anhydride of the formula:##STR7## wherein A, Z and X are each as defined above and Y representsacyl or alkoxycarbonyl can be prepared by the reaction of the compound(II) wherein Y is hydrogen and n is 0 with an acyl halide or an alkylhalocarbonate. Thus, the reaction of 1 molar amount of the compound (II)with a 2 molar amount of an acyl halide (e.g. pivaloyl chloride) or analkyl halocarbonate (e.g. ethyl chlorocarbonate, isobutylchlorocarbonate) in the presence of a 2 molar amount of a basicsubstance may afford the compound (IV) in an excellent yield. (Theprocess using the thus obtained mixed acid anhydride (IV) as thereactant will be referred to as the "mixed anhydride process".)

Another type of the reactive derivative is the compound of the formula:##STR8## wherein A, Z and X are each as defined above, which may beprepared by the reaction of 1 molar amount of the compound (II) whereinY is hydrogen and n is 0 with 1 molar amount of phosgene in the presenceof a 2 molar amount of a basic substance. The similar type of thereactive derivative may be also prepared by the use of thionyl chloride,phosphorus trichloride or the like in place of phosgene. (The processusing the above cyclic compound (V) or any similar compound thereto asthe reactant will be referred to as the "phosgene process".)

Examples of the basic substance in the acid reactions are an inorganicbase (e.g. sodium hydroxide, potassium hydroxide) and an organic base(e.g. triethylamine, pyridine, dimethylaniline, lutidine,N-methylmorpholine and N-methylpiperidine).

Since the reactive derivatives described above are usually very reactiveand unstable to isolate, they may be used in the form of the reactionmixture for the reaction with the compound (III) wherein n is 0.

The derivatives of the compound (III) may be the alkali metal salts(e.g. sodium, potassium salts), the alkaline earth metal salts (e.g.calcium, barium salts), the organic base salts (e.g. trimethylamine,triethylamine salts), the organic sulfonic acid salts (e.g.toluenesulfonic acid, naphthalenesulfonic acid,tetrahydronaphthalenesulfonic acid salts), the esters, the N-substitutedderivatives, etc.

Specific examples of the derivatives of the compound (III) wherein n is0 include the following compounds: ##STR9## wherein R₁, R₂ and R₃ areeach as defined above and R₄, R₅, R₆, R₇ and R₈ are each a lower alkylgroup.

These esters can be advantageously used in the coupling reaction,because of their higher solubility in an ordinary solvent to be used asthe reaction medium and of their higher reactivity with the compound(II) than those of the corresponding free acids.

Further examples of the ester unit in the esters of the compound (III)are as follows: toluenesulfonylethyl ester, p-nitrobenzyl ester, benzylester, phenacyl ester, diphenylmethyl ester, substituted diphenylmethylester, trityl ester, benzoyloxymethyl ester, lower alkanoyloxymethylester, dimethylmethyleneamino ester, p-nitrophenyl ester,methylsulfonylphenyl ester, methylthiophenyl ester, t-butyl ester,3,5-di-t-butyl-4-hydroxybenzyl ester, trichloroethyl ester etc. Theseester units are all conventionally employed as a group protecting acarboxylic acid radical in the related art field.

The esters which can be prepared commercially from penicillin-G areparticularly preferable. Examples of the preparation for such esters areillustratively shown in the following scheme: ##STR10## wherein R₁, R₂and R₃ are each as defined above and -COOE means an ester part.

The above illustrated E ester of the compound (III) can be employed forthe coupling reaction in the form of a salt with an organic or inorganicacid. Examples of the organic or inorganic acid part in such salt aretoluenesulfonic acid, naphthalenesulfonic acid, tetralinesulfonic acid,hydrochloric acid, etc.

The coupling reaction of the compound (II) or its reactive derivativewith the compound (III) or its derivative is usually carried out at atemperature below about 80° C, e.g. at a temperature of -50° to 80° C,but this is not limitative.

The coupling reaction is normally effected in the presence of an inertsolvent. As the inert solvent, there may be used a polar solvent (e.g.dichloromethane, chloroform, acetone, tetrahydrofuran, dioxane,acetonitrile, methylisobutylketone, ethanol, dimethylformamide) or anon-polar solvent (e.g. benzene, toluene, petroleum ether, n-hexane).Water or a water-containing organic solvent is also utilizable dependingon the type of starting materials used.

In case that the compound (II) is subjected to coupling in a free formor a salt form, the reaction may be carried out preferably in thepresence of a conventional coupling reagent (e.g.N,N'-dicyclohexylcarbodiimide, diphenyl phosphorous acid).

In the compound (II), the substituent --X--Y may represent either a freehydroxyl or sulfhydryl group or a protected hydroxyl or sulfhydrylgroup. When the compound (II) wherein the substituent --X--Y representsa protected hydroxyl or sulfhydryl group is employed in the couplingreaction, the penicillin (I) wherein Y is hydrogen may be often obtainedas the result of simultaneous elimination of the protective group. Whenthe protective group is not eliminated in the course of the couplingreaction, it may be eliminated thereafter by a conventional procedureunder such a mild condition that the opening of the lactam ring in thepenicillin nucleus is not caused. The elimination of the protectivegroup can be accomplished, for instance, by treating the product in thecoupling reaction with an inorganic or organic basic substance (e.g.sodium carbonate, potassium carbonate, sodium hydroxide, aqueous ammoniasolution, triethylamine, methylamine, dimethylamine, diethylamine,morpholine, piperidine, potassium acetate, sodium acetate, potassium2-ethylhexanoate). In such treatment, the penicillin (I) wherein Y ishydrogen can be obtained even in an acidic condition, but the protectivegroup is more smoothly eliminated by treatment under a basic condition.

In case that the compound (II) wherein n is 0 and Y is loweralkoxycarbonyl or lower alkanoyl and the compound (III) wherein n is Oand R₁ is hydroxyl are subjected to coupling, there may be sometimesobtained the penicillin (I) wherein Y is hydrogen and R₁ is loweralkoxycarbonyl or lower alkanoyl. Further, the coupling reaction of thecompound (III) wherein n is O and R₁ is protected hydroxyl (e.g.ethoxycarbonyloxy, benzyloxycarbonyloxy) with the compound (II) whereinn is O and Y is lower alkoxycarbonyl or lower alkanoyl may afford thepenicillin (I) wherein Y is lower alkoxycarbonyl or lower alkanoyl andR₁ is protected hydroxyl.

When the compound (II) wherein Y is hydrogen is subjected to coupling,it may be favorably employed, for instance, in the form of the reactiveester or the acid halide on the carboxyl group whereby the penicilline(I) wherein Y is hydrogen is obtainable as the product.

The penicillin (I) wherein the ring A is substituted with an amino groupmay be produced from the corresponding penicillin (I) wherein the ring Ais substituted with a nitro group or a protected amino group. Forexample, the reduction of the penicillin (I) wherein the ring A bears anitro group or a benzyloxycarbonylamino group under such a mildcondition that the opening of the lactam ring in the pencillin nucleusis not caused gives the penicillin (I) wherein the ring A bears an aminogroup. Further, for example, the hydrolysis of the penicillin (I)wherein the ring A bears a protected amino group (e.g. enamine) under amild condition as above results in the elimination of the protectivegroup to give the penicillin (I) wherein the ring a bears an aminogroup.

Alternatively, the penicillin (I) having an amino group on the ring amay be produced by the said coupling reaction wherein the compound (II)is used in the form of the acid halide on the carboxyl group.

When any protective group is present in the product of the couplingreaction, it may be eliminated by a per se conventional procedure suchas catalytic reduction or hydrolysis, favorably under a mild condition.

The production of the penicillin (I) may be identified by thin layerchromatography, iodometry, infrared absorption spectrum and so on. Thecharacteristic infrared absorption due to the lactam ring is at 1750 -1800 cm⁻ ¹. A particularly effective identifying method is NMR analysis,since the signals attributed to the proton Ha of the amide bond in thefollowing structure for the penicillin (I) appear in a very low-field,which is due to the presence of the substituent --X--Y: ##STR11##

When measured in hexadeuterodimethylsulfoxide at 60 MHz using anNMR-spectral instrument, Ha and Hb signals in case of Y being a loweralkanoyl group or a lower alkoxycarbonyl group appear respectively at540 - 570 Hz and at 630 - 680 Hz. In case of Y being a hydrogen atom, Haand Hb signals appear respectively at 650 - 690 Hz and at 540 - 570 Hz.Ha and Hb signals in case of Y being a hydrogen atom appear in a lowerfield than those of Y being a lower alkanoyl group or a loweralkoxycarbonyl group.

The produced penicillin (I) may be, if desired, converted into itsnon-toxic pharmaceutically acceptable salt in a per se conventionalmanner.

Still, the compound (II) wherein n is l can be prepared easily by aconventional procedure, for instance, by reacting the reactivederivative of the compound (II) wherein n is O with an amino acid of theformula: ##STR12## wherein R₁, R₂ and R₃ are each as defined above, orits ester in water or an organic solvent in the presence of a basicsubstance.

The amino acid may be any of the DL-, D- and L-configurations. Theesters may be, for example, trialkylsilyl ester, lower alkyl ester,p-nitrophenyl ester, benzyl ester, phenylthiophenyl ester,N-hydroxysuccinimide ester, etc. These esters may be derived from thecorresponding acid chlorides or prepared by any other conventionalmethod. As the organic solvent, a polar or non-polar organic solvent(e.g. dioxane, tetrahydrofuran, dichloromethane, benzene,dimethylformamide, dimethylsulfoxide) is utilizable. Examples of thebasic substance are sodium hydroxide, potassium hydroxide,triethylamine, N-methylmorpholine, dimethylaniline, etc.

When the ester is employed in the above reaction, the protective groupof the resulting N-acylamino acid ester may be eliminated by aconventional procedure to give the compound (II) wherein n l. Whendesired, the obtained active ester salt of the compound (II) wherein n lmay be employed in the coupling reaction with the compound (III) whereinn is l to produce the penicillin (I).

The present invention is illustrated more precisely by the followingExamples but those are not intended to limit the scope of the presentinvention.

EXAMPLE 1

Preparation ofD-α-(4-hydroxy-1,5-naphthyridine-3-carboxyamido)-p-hydroxybenzylpenicillin:-##STR13##

One hundred ml of dichloromethane and 2.2 g of triethylamine were addedto 3.87 g of D-α-amino-p-hydroxybenzylpenicillin sodium salt, and 2.2 gof trimethylchlorosilane were dropwise added thereto. To the resultingdichloromethane solution of the pencillin trimethylsilyl ester, therewere added 2.2 g of triethylamine and then 2.45 g of powdered4-hydroxy-1,5-naphthyridine-3-carbonyl chloride hydrochloride at 0° to5° C. The resultant mixture was stirred at 0° 5° C overnight. Thereaction mixture was adjusted to pH 2 with a 11.2% solution of hydrogenchloride in dioxane while cooling with ice. The solvent was evaporatedunder reduced pressure, and 100 ml of ice water were added to theresidue. The precipitated crystals were collected by filtration, washedwith water and dissolved in an aqueous solution of sodium bicarbonate.The insoluble materials were separated by filtration, and the filtratewas adjusted to pH 2 with N hydrochloric acid while cooling with ice.The precipitated crystals were collected by filtration and dried underreduced pressure to give 3.1 g of the objective compound as crystals.Purity (determined by iodometry), 86.5%.

4-Hydroxy-1,5-naphthyridine-3-carbonyl chloride hydrochloride used asthe starting material in the above Example was prepared as follows:

4-Hydroxy-1,5-naphthyridine-3-carboxylic acid was added to a mixture ofbenzene, 0.73 g of dimethylformamide and 1.56 g of thionyl chloride, andthe resulting mixture was stirred at 40° to 85° C for 3 hours. Theprecipitated crystals were collected by filtration, washed with benzeneand dried under reduced pressure to give 2.37 g of4-hydroxy-1,5-naphthyridine-3-carbonyl chloride hydrochloride ascrystals. Purity (determined by NMR analysis as the ester form obtainedby further alcoholysis), 96.3%.

EXAMPLE 2

Preparation ofDL-α-(4-hydroxy-1,5-naphthyridine-3-carboxamido)-p-hydroxybenzylpenicillin:-##STR14##

a. To the mixture of 2.22 g of DL-α-amino-p-hydroxybenzylpenicillintriethylamine salt and 15 ml of dimethylformamide, there were added 1.3g of triethylamine and then 1.08 g of powdered4-hydroxyl-1,5-naphthyridine-3-carbonyl chloride hydrochloride whilecooling with ice, and the resulting mixture was stirred for 4 hours. Theprecipitate was collected by filtration, washed with dimethylformamideand dichloromethane in order to give 1.2 g ofDL-α-(4-hydroxy-1,5-naphthyridine-3-carboxamido)-p-hydroxybenzylpenicillintriethylamine salt as brown crystals. IR ν_(C) _(=O) 1775 cm⁻ ¹, 1660cm⁻ ¹.

The filtrate and the washing solvents were combined together, and 100 mlof ether were added thereto. The insoluble material was collected byfiltration and treated with 50 ml of dichloromethane. The obtainedcrystals were suspended in 30 ml of dichloromethane, adjusted to pH 2with an 11.2% solution of hydrogen chloride in dioxane while coolingwith ice and concentrated under reduced pressure. The residue wasadmixed with 30 ml of cold water, collected by filtration, washed withwater and dried under reduced pressure to give 1.1 g ofDL-α-(4-hydroxy-1,5-naphthyridine-3-carboxyamido)-p-hydroxybenzylpenicillinas brown crystals.

To 0.93 g ofDL-α-(4-hydroxy-1,5-naphthyridine-3-carboxamido)-p-hydroxybenzylpenicillintriethylamine salt as obtained above, there were added 6 ml ofdimethylformamide and then 0.792 g of 50% solution of potassium2-ethylhexanoate in n-butanol, and the resultant mixture was stirred atroom temperature for 20 minutes. After removal of insolube materials byfiltration, the filtrate was added to 25 ml of acetone heated at 40 to50° C while stirring. The precipitated crystals were immediatelycollected by filtration, washed with acetone and dried under reducedpressure to giveDL-α-(4-hydroxy-1,5-naphthyridine-3-carboxamido)-p-phydroxybenzylpenicillinpotassium salt. Purity (determined by iodometry), 87%.

In the same procedure as above,DL-α-(4-hydroxy-1,5-naphthyridine-3-carboxamido)-p-hydroxybenzylpenicillinpotassium salt was prepared fromDL-α-(4-hydroxy-1,5-naphthyridine-3-carboxamido)-p-hydroxybenzylpenicillinas obtained above.

b. To a suspension of 0.5 g ofDL-α-(4-hydroxy-1,5-naphthyridine-3-carboxamido)-p-hydroxyphenylaceticacid in 15 ml of anhydrous dimethylsulfoxide, 0.27 g ofN,N'-carbonyldiimidazole was added while stirring and cooling with ice.After 30 minutes, 0.3 g of powdered 6-aminopenicillanic acid was addedthereto, and the resulting mixture was stirred for 4 hours. The reactionmixture was filtered, and the filtrate was dropwise added to 200 ml ofacetone while stirring. The precipitate was collected by filtration anddried under reduced pressure. The obtained crude penicillin wasdissolved in dimethylformamide and, after removal of insoluble materialsby filtration, 50% solution of potassium 2-ethylhexanoate in n-butanolwas added. The resultant mixture was added dropwise to 40 ml of acetoneunder reflux while stirring. After 10 minutes, the precipitated crystalswere collected by filtration to giveDL-α-(4-hydroxy-1,5-naphthyridine-3-carboxamido)-p-hydroxybenzylpenicillinpotassium salt. Purity (determined by iodometry), 78%.

Dl-α-(4-Hydroxy-1,5-naphthyridine-3-carboxamido)-p-hydroxyphenylaceticacid used as the starting material in the part (b) of the above Examplewas prepared as follows:

To a mixture of 1.35 g of DL-2-p-hydroxyphenylglycine, 50 ml ofanhydrous dichloromethane and 4.05 g of triethylamine, 2.62 g oftrimethylchlorosilane were added, and the resulting mixture was refluxedfor 20 minutes. After the addition of 2.1 g of4-hydroxy-1,5-naphthyridine-3-carbonyl chloride hydrochloride, thereaction was effected while cooling with ice for 4 hours. The reactionmixture was concentrated under reduced pressure, 50 ml of water wereadded to the residue, and the resulting mixture was adjusted to pH 1with 4N hydrochloric acid. The precipitated crystals were collected byfiltration, suspended in 50 ml of water and potassium carbonate wasadded thereto. After removal of insoluble materials by filtratior, thefiltrate was adjusted to pH 1. The precipitated crystals were collectedby filtration, washed with water and dried to give 2.1 g ofDL-α-(4-hydroxy-1,5-naphthyridine-3-carboxamido)-p-hydroxyphenylaceticacid. M.P. 310° to 320° C (decomp.).

EXAMPLE 3

Preparation ofDL-α-(4-hydroxy-6-dimethylamino-1,5-naphthyridine-3-carboxamido)-p-ethoxycarbonyloxybenzylpenicillin:-##STR15##

In a mixture of 80 ml of dichloromethane and 1.01 g of triethylamine,1.17 g of 4-hydroxy-6-dimethylamino-1,5-naphthyridine-3-carboxylic acidwere suspended, and 1.09 g of ethyl chlorocarbonate were added theretoat -20 to -25° C. After the reaction is effected for 1 hour, a mixtureof 1.85 g of powdered DL-α-amino-p-hydroxybenzylpenicillin triethylaminesalt, 30 ml of dichloromethane and 0.25 g of triethylamine was pouredtherein, and the resulting mixture was stirred at -25° C for 5 hours.The reaction mixture was admixed with a solution of 1.26 g of sodiumbicarbonate in 300 ml of water and shaken with 300 ml of ethyl acetate.The aqueous layer was separated and adjusted to pH 3.5 with Nhydrochloric acid. The precipitated crystals were collected byfiltration, washed with water and dried under reduced pressure to give2.26 g ofDL-α-(4-hydroxy-6-dimethylamino-1,5-naphthyridine-3-carboxamido)-p-ethoxycarbonyloxybenzylpenicillin.

EXAMPLE 4

Preparation ofD-α-(4-hydroxyquinoline-3-carboxamido)-p-pivaloyloxybenzylpenicillin:-##STR16##

To a mixture of 1.88 g of 4-hydroxyquinoline-3-carboxylic acid, 60 ml ofdichloromethane and 2.02 g of triethylamine while cooling with ice andstirring, 2.5 g of pivaloyl chloride were added dropwise for 1 hour. Tothe resulting mixture, 4.67 g of powderedD-α-amino-p-hydroxybenzylpenicillin triethylamine salt were added, andthe reaction was effected. The reaction mixture was admixed with an11.2% solution of hydrogen chloride in dioxane to make pH 2 andconcentrated under reduced pressure. The residue was suspended in 60 mlof ice water, and the precipitate was collected by filtration and driedunder reduced pressure to giveD-α-(4-hydroxyquinoline-3-carboxamido)-p-pivaloyloxybenzylpenicillin ascrystals. Purity (determined by iodometry), 78.3%.

EXAMPLE 5

Preparation ofD-α-(4-ethoxycarbonyloxy-1,5-naphthyridine-3-carboxamido)-p-benzyloxycarbonyloxybenzylpenicillin:##STR17##

To a mixture of 0.95 g of powdered 4-hydroxy-1,5-naphthyridine, 20 ml ofdichloromethane and 1.1 g of triethylamine maintained at 0° to 5° C,0.55 g of ethyl chlorocarbonate was added, and the resultant mixture wasstirred for 30 minutes. Then, 2.9 g ofD-α-amino-p-benzyloxycarbonyloxybenzylpenicillin triethylamine salt wereadded thereto, and the reaction was effected for 7 hours. The reactionmixture was adjusted to pH 2 at 0° C with an 11.2% solution of hydrogenchloride in dioxane and concentrated under reduced pressure. The residuewas admixed with 20 ml of ice water and adjusted to pH 2. Theprecipitated crystals were collected by filtration and washed with waterto giveD-α-(4-ethoxycarbonyloxy-1,5-napthyridine-3-carboxamido)-p-benzyloxycarbonyloxybenzylpenicillin.Purity (determined by iodometry, 76%.

EXAMPLE 6

Preparation ofDL-α-(4-hydroxyquinoline-3-carboxamido)-p-hydroxybenzylpenicillin:##STR18##

To a mixture of 1.88 g of 4-hydroxyquinoline-3-carboxylic acid, 60 ml ofdichloromethane and 2.2 g of triethylamine kept at -20° to -25° C, 9.9 gof 10% solution of phosgene in dichloromethane were dropwise added, andthe resultant mixture was stirred for 30 minutes. Then, 3.9 g ofpowdered DL-α-amino-p-hydroxybenzylpenicillin sodium salt were addedthereto, and the reaction was effected at the same temperature as abovefor 5 hours and at 0° C for 1 hour. The reaction mixture was treated asin Example 4 to give 3.8 g ofDL-α-(4-hydroxyquinoline-3-carboxamido)-p-hydroxybenzylpenicillin.Purity (determined by iodometry, 81.5%.

EXAMPLE 7

Preparation ofD-α-(4-hydroxy-7-methyl-1,8-naphthyridine-3-carboxamido)-p-hydroxybenzylpenicillin:-##STR19##

To a solution of 0.5 g of D-α-amino-p-hydroxybenzylpenicillin sodiumsalt in 50 ml of dimethylformamide, 0.387 g of powdered4-hydroxy-7-methyl-1,8-naphthyridine-3-carboxylic acid p-nitrophenylester was added, and the resulting mixture was stirred at roomtemperature for 10 hours. After removal of insoluble materials befiltration, 200 ml of ether were added to the filtrate. The precipitatewas collected by filtration, dissolved in 10 ml of water and adjusted topH 2 with dilute hydrochloric acid while cooling with ice. Theprecipitated crystals were collected by filtration and dried underreduced pressure to giveD-α-(4-hydroxy-7-methyl-1,8-naphthyridine-3-carboxamido)-p-hydroxybenzylpenicillin.Purity (determined by iodometry), 82.7%.

4-Hydroxy-7-methyl-1,8-naphthyridine-3-carboxylic acid p-nitrophenylester used as the starting material in the above Example was prepared asfollows:

A mixture of 1 g of 4-hydroxy-7-methyl-1,8-naphthyridine-3-carboxylicacid and 25 ml of anhydrous pyridine was heated on an oil bath at 50° to60° C while stirring. Then, 1.38 g of p-nitrophenyl trifluoroacetatewere added thereto and, after the heating was interrupted, stirring wascarried out for 2 hours. The precipitated crystals were collected byfiltration and washed with ethanol to give4-hydroxy-7-methyl-1,8-naphthyridine-3-carboxylic acid p-nitrophenylester as pale yellow crystalline powder melting at 311 to 313° C(decomp.). Yield, 1.2 g. IR ν_(C) _(=O) 1700 cm⁻ ¹ ; ν_(NO).sbsb.2 1520cm⁻ ¹, 1345 cm⁻ ¹.

EXAMPLE 8

Preparation ofD-α-(4-hydroxycinnoline-3-carboxamido)-p-hydroxybenzylpenicillin:-##STR20##

In 30 ml of anhydrous dimethylformamide, 0.95 g of4-hydroxycinnoline-3-carboxylic acid was dissolved at room temperature,and 0.89 g of carbonyldiimidazole was added thereto. After 30 minutes, asolution of 2.34 g of D-α-amino-p-hydroxybenzylpenicillin triethylaminesalt in 20 ml of dimethylformamide was added to the resulting mixture,and stirring was continued at room temperature for 6 hours. After theaddition of 1.82 g of a 50 % solution of potassium 2-ethylhexanoate inn-butanol and then ether, the resultant mixture was filtered. Thecollected substance was dissolved in water and adjusted to pH 2 with Nhydrochloric acid while cooling with ice. The precipitate was collectedby the filtration and dried over phosphorus pentoxide under reducedpressure to give 1.7 g ofD-α-(4-hydroxycinnoline-3-carboxamido)-p-hydroxybenzylpenicillin. Thisproduct was suspended in 50 ml of acetane, 1.4 g of 50 % solution ofpotassium 2-ethylhexanoate in n-butanol were added to the suspension,and refluxing was continued for 5 minutes. The precipitated crystalswere collected by filtration while hot to give 1.6 g ofD-α-(4-hydroxycinnoline-3-carboxamido)-p-hydroxybenzylpenicillinpotassium salt. Purity (determined by iodometry), 86.2%.

EXAMPLE 9

Preparation ofD-α-(4-hydroxy-1,5-naphthyridine-3-carboxamido)-p-ethoxycarbonyloxybenzylpenicillin:-##STR21##

To a mixture of 0.95 g of powdered4-hydroxy-1,5-naphthyridine-3-carboxylic acid, 25 ml of dichloromethaneand 1.1 g of triethylamine, 1.1 g of ethyl chlorocarbonate were added at0° C, and the resultant mixture was stirred at the same temperature for40 minutes. After the addition of 2.9 g ofD-α-amino-p-hydroxybenzylpenicillin 3', 5'-di-t-butyl-4'-hydroxybenzylester (prepared from penicillin G according to a conventionalprocedure), the reaction was effected at the same temperature as aboveovernight. The solvent was removed from the reaction mixture bydistillation under reduced pressure, and cold water was added thereto.The insoluble material was collected by filtration and dried underreduced pressure. The thus obtained product was admixed with 30 ml ofanhydrous dimethylformamide and 1.8 g of potassium 2-ethylhexanoate, andthe resulting mixture was dropwise added to 100 ml of acetone. Theprecipitate was collected by filtration and washed with acetone to giveD-α-(4-hydroxy-1,5-naphthyridine-3-carboxamido)-p-ethoxycarbonyloxybenzylpenicillinpotassium salt. Purity (determined by iodometry), 78.7%.

EXAMPLES 10-41

In the same procedure as above, there were produced the penicillins (I)as shown in Table 1.

                                      Table 1                                     __________________________________________________________________________     ##STR22##                                                                    Exam-                                                                         ple                                        Purity                             No. L                   R.sub.1  R.sub.2                                                                          R.sub.3                                                                          M   (%)                                __________________________________________________________________________    10                                                                                 ##STR23##          p-OCOOC.sub.2 H.sub.5                                                                  H  H  H   81.3                               11                                                                                 ##STR24##          p-OH     H  H  K   83.0                               12                                                                                 ##STR25##          p-OCOOC.sub.2 H.sub.5                                                                  H  H  H   88.3                               13                                                                                 ##STR26##          m-OH     H  H  Na  85.5                               14                                                                                 ##STR27##          p-OH     m-Cl                                                                             H  K   84.9                               15                                                                                 ##STR28##          p-OCOOC.sub.2 H.sub.5                                                                  H  H  H   88.1                               16                                                                                 ##STR29##          m-OH     H  H  H (tri- ethyl- amine salt                                                         89.0                               17                                                                                 ##STR30##          p-OCOOC.sub.2 H.sub.5                                                                  H  H  H   89.5                               18                                                                                 ##STR31##          p-OH     H  H  K   88.0                               19                                                                                 ##STR32##          p-OH     H  H  K   85.4                               20                                                                                 ##STR33##          p-OCOOC.sub.2 H.sub.5                                                                  H  H  H   81.5                               21                                                                                 ##STR34##                                                                     ##STR35##          p-OH     H  H  K   84.2                               22                                                                                 ##STR36##          p-OH     H  H  K   78.1                               23                                                                                 ##STR37##          p-OCOOC.sub.2 H.sub.5                                                                  H  H  H   83.0                               24                                                                                 ##STR38##          p-OH     H  H  K   85.7                               25                                                                                 ##STR39##          p-OCOOC.sub.2 H.sub.5                                                                  m-Cl                                                                             H  H   89.4                               26                                                                                 ##STR40##          m-OH     H  H  K   87.0                               27                                                                                 ##STR41##          p-OH     H  H  H   80.0                               28                                                                                 ##STR42##          p-OH     H  H  Na  83.2                               29                                                                                 ##STR43##          p-OH     H  H  K   87.5                               30                                                                                 ##STR44##                                                                                         ##STR45##                                                                             H  H  H   77.3                               31                                                                                 ##STR46##          p-OH     H  H  K   81.0                               32                                                                                 ##STR47##          p-OH     m-Cl                                                                             H  K   87.8                               33                                                                                 ##STR48##          p-OCOOC.sub.2 H.sub.5                                                                  H  H  H   88.0                               34                                                                                 ##STR49##          p-OH     H  H  K   87.3                               35                                                                                 ##STR50##          p-OH     H  H  K   82.1                               36                                                                                 ##STR51##          p-OCOOC.sub.2 H.sub.5                                                                  H  H  H   80.1                               37                                                                                 ##STR52##          p-OCOOC.sub.2 H.sub.5                                                                  H  H  H   90.2                               38                                                                                 ##STR53##          p-OCOOC.sub.2 H.sub.5                                                                  H  H  H   75.9                               39                                                                                 ##STR54##          p-OCOOC.sub.2 H.sub.5                                                                  H  H  H   88.1                               40                                                                                 ##STR55##          p-OH     m-Cl                                                                             m'-Cl                                                                            K   77.3                               41                                                                                 ##STR56##          p-OH     H  H  H   82.5                               42                                                                                 ##STR57##          p-OCOOC.sub.2 H.sub.5                                                                  H  H  H   85.1                               43                                                                                 ##STR58##          p-OH     H  H  K   88.3                               __________________________________________________________________________     Note:                                                                         1) The penicillins (I) in Example Nos. 16, 26 and 40 are in the               DL-configuration, and the others are all in the D-configuration.              2) The purity was determined by iodometry.                               

EXAMPLE 44

Preparation ofD-α-(4-hydroxy-1,5-naphthyridine-3-carboxamido)-p-hydroxybenzylpenicillin:##STR59##

To a mixture of 3.9 g of D-α-amino-p-hydroxybenzylpenicillin, 40 ml ofdimethylformamide and 1.88 g of triethylamine, 2.8 g of theN-hydroxysuccinimide ester of 4-hydroxy-1,5-naphthyridine-3-carboxylicacid and 20 ml of dimethylformamide were added, and stirring waseffected at room temperature for 5 hours. The resulting mixture waspoured into 160 ml of acetone and stirred for 20 minutes. Then, thereaction mixture was filtered, and the collected substance was washedwith acetone and dichloromethane in order and dried at 30 to 40° C underreduced pressure to give 4.9 g ofD-α-(4-hydroxy-1,5-naphthyridine-3-carboxamido)-(-)p-hydroxybenzylpenicillintriethylamine salt. A mixture of 4.3 g of the thus preparedtriethylamine salt, 30 ml of dimethylformamide and 1.34 g of sodium2-ethylhexanoate was dropwise added to 150 ml of acetone. Theprecipitated crystals were collected by filtration, washed with acetoneand dried under reduced pressure to give 3.5 g ofD-α-(4-hydroxy-1,5-naphthyridine-3-carboxamido-p-hydroxybenzylpenicillinsodium salt Purity (determined by iodometry), 910000040039133 %.

N-Hydroxysuccinimide ester of 4-hydroxy-1,5- naphthyridine-3-carboxylicacid used as the starting material in the above Example was prepared asfollows:

To a mixture of 7.6 g of 4-hydroxy-1,5-naphthyridine-3-carboxylic acid,5.06 g of N-hydroxypuccinimide and 150 ml of dimethylformamide, therewere dropwise added 5.72 g of thionyl chloride, and the resultingmixture was stirred at room temperature for 16 hours. Then, 8.23 g ofpyridine were dropwise added thereto at a temperature below 10° C, andstirring was continued at room temperature for 4 hours. The precipitatewas collected by filtration, washed with dimethylformamide and acetonein order and dried to give 11.2 g of N-hydroxysuccinimide ester of the4-hydroxy-1,5-naphthyridine-3-carboxylic acid. M.P. 258 to 260° C(decomp.).

When determined according to the agar dilution method, the penicillins(I) afford the minimal inhibitory concentrations against testmicroorganisms as shown in Table 2.

                                      Table 2                                     __________________________________________________________________________    Minimal inhibitory concentration(μg/ml)                                                                Klebsie-                                                                           Pseudo-                                          Staphylo-                                                                           Escheri-                                                                            lla   monas                                                   Exam-                                                                             coccus                                                                              chia  Proteus                                                                             Proteus                                                                             pneumo-                                                                            aerugi-                                      ple aureus                                                                              coli  miravilis                                                                           vulgaris                                                                            niae nosa                                         No. 209P  NIHJ  GN2425                                                                              HV19  PC1602                                                                             104                                          __________________________________________________________________________     1  0.78  1.56  1.56  0.025 12.5 1.56                                          2  1.56  3.13  3.13  0.025 25   1.56                                          3  1.56  1.56  1.56  0.1   3.13 3.13                                          4  0.39  6.25  6.25  0.2   12.5 6.25                                          5  1.56  3.13  3.13  0.05  12.5 3.13                                          6  0.78  6.25  6.25  0.2   25   6.25                                          7  0.78  1.56  3.13  0.05  12.5 1.56                                          8  0.78  12.5  12.5  0.78  25   12.5                                          9  1.56  3.13  3.13  0.05  12.5 3.13                                         10  0.39  3.13  0.78  --    3.13 3.13                                         11  0.78  12.5  3.13  --    0.78 12.5                                         12  0.78  3.13  3.13  0.2   25   6.25                                         13  0.78  6.25  3.13  0.2   25   6.25                                         14  0.78  6.25  6.25  --    25   6.25                                         15  1.56  3.13  6.25  0.1   12.5 3.13                                         16  1.56  3.13  3.13  0.05  25   3.13                                         17  0.78  1.56  1.56  0.1   3.13 1.56                                         18  0.78  1.56  1.56  0.05  12.5 1.56                                         19  0.78  3.13  3.13  0.05  12.5 3.13                                         20  0.78  12.5  6.5   --    3.13 12.5                                         21  0.39  12.5  12.5  --    3.13 12.5                                         22  0.78  1.56  1.56  0.05  12.5 3.13                                         23  1.56  3.13  3.13  0.2   25   3.13                                         24  1.56  1.56  3.13  0.05  12.5 3.13                                         25  0.78  3.13  3.13  0.05  12.5 3.13                                         26  1.56  3.13  3.13  0.05  25   3.13                                         27  0.78  6.25  3.13  0.2   12.5 6.25                                         28  0.78  3.13  3.13  0.05  12.5 3.13                                         29  0.78  12.5  12.5  0.2   25   6.25                                         30  0.78  6.25  3.13  0.05  12.5 6.25                                         31  0.78  3.13  3.13  0.1   12.5 3.13                                         32  0.78  3.13  6.25  0.05  25   3.13                                         33  0.78  3.13  3.13  0.1   12.5 6.25                                         34  0.78  1.56  1.56  0.05  12.5 1.56                                         35  0.78  1.56  1.56  0.025 12.5 1.56                                         36  0.78  3.13  3.13  0.05  25   3.13                                         37  0.78  1.56  1.56  0.025 12.5 1.56                                         38  0.39  6.25  3.13  0.2   6.25 3.13                                         39  0.78  1.56  1.56  0.05  6.25 1.56                                         40  1.56  12.5  6.25  0.2   25   6.25                                         41  1.56  6.25  6.25  0.2   25   3.13                                         42  0.78  3.13  6.25  0.2   12.5 6.25                                         43  0.78  1.56  1.56  0.025 6.25 1.56                                         Com-                                                                          pound                                                                         R   0.78  100   50    0.39  50   50                                           Ampi-                                                                         cillin                                                                            0.1   6.26  1.56  1.56  50   > 200                                        Amoxy-                                                                        cillin                                                                            0.2   12.5  3.13  12.5  > 200                                                                              > 200                                        Car-                                                                          beni-                                                                         cillin                                                                            0.78  12.5  0.78  0.78  > 200                                                                              50-100                                       __________________________________________________________________________    Note: The compound R is described in U.S. Pat. No.                            3,433,784 and has the following chemical structure:                            ##STR60##                                                                

What is claimed is:
 1. A compound of the formula: ##STR61##wherein Z isa nitrogen atom or a methylene group, the ring A is a benzene, pyrazole,thiazole, imidazole, pyrazine, pyridazine or pyrimidine ring when Z is anitrogen atom or a methylene group, or a pyridine ring when Z is anitrogen atom, the ring A being unsubstituted on being substituted withone on more substituents selected from the group consisting of loweralkyl, lower alkoxy, lower alkylthio, lower haloalkyl, loweralkylenedioxy, halogen, hydroxyl, nitro, amino, loweralkoxycarbonylamino, lower alkylamino, di(lower)alkylamino and loweralkanoylamino, R₁ is hydroxyl, lower alkanoxyloxy, loweralkoxycarbonyloxy or benzyloxycarbonyloxy, R₂ and R₃ are each hydrogenor halogen, X is oxygen or sulfur and Y is hydrogen, loweralkoxycarbonyl or lower alkanoyl, the Y-X- group being present on thecarbon atom adjacent to the carbon atom to which the6-(α-aminoacylamido)penicillanic acid moiety is linked, and non-toxicpharmaceutically acceptable salts thereof.
 2. The compound according toclaim 1, wherein Y is a hydrogen atom.
 3. The compound according toclaim 1, wherein R₁ is a hydroxyl group, a C_(1-C) ₆ alkoxycarbonyloxygroup or a benzyloxycarbonyl group and R₂ and R₃ are each a hydrogenatom or a chlorine atom.
 4. The compound according to claim 1, whereinthe ring A is an unsubstituted benzene ring or a benzene ring having oneor more substituents selected from the group consisting of chlorine,trifluoromethyl, nitro, benzyloxycarbonyl amino and methylenedioxy. 5.The compound according to claim 4, wherein R₁ is a hydroxyl group or anethoxycarbonyloxy group and R₂ and R₃ are each a hydrogen atom or achlorine atom.
 6. A compound of the formula: ##STR62##wherein the ring Ais a benzene, pyrazole, thiazole, imidazole, pyrazine, pyridazine orpyrimidine ring, said ring being unsubstituted or substituted with oneor more substituents selected from the group consisting of C_(1-C) ₈alkyl, C_(1-C) ₈ alkoxy, C_(1-C) ₈ alkylthio, C_(1-C) ₈ haloalkyl,C_(1-C) ₃ alkylenedioxy, halogen, nitro, free or protected amino,C_(1-C) ₈ alkylamino, di(C_(1-C) ₈)alkylamino and C_(1-C) ₉alkanoylamino, R₁ is a free or protected hydroxyl group and R₂ and R₃are each a hydrogen atom or a halogen atom, and non-toxic,pharmaceutically acceptable salts thereof.
 7. A compound of the formula:##STR63##wherein R₁ is a free or protected hydroxyl group and R₂ and R₃are each a hydrogen atom or a halogen atom.
 8. A compound of theformula: ##STR64## 9.D-α-(7-Chloro-4-hydroxyquinoline-3-carboxamido)-p-ethoxycarbonyloxybenzylpenicillinand non-toxic, pharmaceutically acceptable salts thereof. 10.D-α-(4-Ethoxycarbonyloxyquinoline-3-carboxamido)-p-ethoxycarboxyloxybenzylpenicillinand non-toxic, pharmaceutically acceptable salts thereof. 11.D-α-(4-Hydroxyquinoline-3-carboxamido)-m-chloro-p-hydroxybenzylpenicillinand non-toxic, pharmaceutically acceptable salts thereof. 12.D-α-{8-Hydroxypyrido[3,2-d]pyrimidine-7-carboxamido}-p-hydroxybenzylpenicillinand non-toxic, pharmaceutically acceptable salts thereof. 13.D-α-{2,4-Dimethyl-8-hydroxypyrido[3,2-d]-pyrimidine-7-carboxamido}-p-hydroxybenzylpenicillinand non-toxic, pharmaceutically acceptable salts thereof.